To obtain information about protein requirements and utilization, practical and basic methodology will be used to investigate dietary crude protein requirements, rumen bypass of potentially limiting nutrients, mammary gland amino acid metabolism and gluconeogenesis in ruminants. Studies will be specifically designed to determine the dietary crude protein requirements of lactating cows and the effect on animal performance of substituting urea for protein in the diet of lactating cows. Since our previous work has shown that rumen bypass of casein will increase both milk and milk protein yields, other more economical sources of protein as well as mixtures of amino acids will be used in rumen bypass studies to determine their effects on lactational performance. Our previous work indicates that valine, isoleucine and leucine are extracted by the mammary gland in larger quantities than their output in milk, thus, their metabolism in the mammary gland will be investigated using bovine mammary tissue slices. Ruminants unlike nonruminants must rely almost totally on gluconeogenesis for their supply of glucose. Ruminants also absorb large amounts of ammonia into the portal blood and ammonia is known to cause profound alterations in intermediary carbohydrate metabolism. Therefore in vitro liver tissue slice incubations and in vivo glucose entry rate measurements will be used to determine the influence of ammonia on glucose synthesis in ruminants. These studies will provide an insight for dietary crude protein requirements of lactating cows, a knowledge of which nutrients improve animal performance when they bypass rumen fermentation and a comprehension of intracellular regulation of nitrogen and carbohydrate metabolism. Through a better understanding of nutrient requirements and utilization, feed efficiency and total food output by ruminant animals should be improved, resulting in an improved nutrition and health of the world's population.